Development and validation of a novel hydrolysis probe real-time polymerase chain reaction for agamid adenovirus 1 in the central bearded dragon (Pogona vitticeps)

Abstract

Adenoviruses are nonenveloped DNA viruses that range from 60 to 100 nm in diameter. Infections with adenoviruses are well documented, and have been reported in all of the bony vertebrates, including squamates, from which the genus Atadenovirus likely originated. 3,5,19 Currently, there are 5 accepted genera of adenoviruses within the family Adenoviridae: Atadenovirus (predominantly in squamate hosts), Aviadenovirus (in avian hosts), Ichtadenovirus (isolated from a white sturgeon), Mastadenovirus (in mammalian hosts), and Siadenovirus (found in chelonian, amphibian, and avian species). Additionally, a sixth genus has been proposed (Testadenovirus), whose members were isolated from hosts in the order Testudines.2 Many reptilian adenoviruses appear to have coevolved with the species they infect, leading to a fair amount of epidemiological species specificity. [17][18][19] Bearded dragons (Pogona sp.) are agamid lizards native to Australia that have become an increasingly popular pet and are frequently found in zoological collections. Although exportation of these animals from Australia is illegal, a successful breeding industry has led to numerous animals in captivity. The most common species, the central bearded dragon (Pogona vitticeps), is represented by more than 700 individuals in 258 zoological institutions worldwide (International Species Information System. Species holdings. 2014. https://zims.isis.org). In general, bearded dragons make good pets, and typically thrive in captivity, although a number of diseases have been documented.Agamid adenovirus 1 (AgAdV-1) appears to be a highly prevalent disease of concern in Pogona species. Adenoviral disease was first reported in 1982, 10 and there have been numerous documented cases of AgAdV-1 infections in bearded dragons. 4,9,11 In 2004, the first specific characterization of AgAdV-1 was reported, 19 and it appears to be widespread in P. vitticeps, having been identified in numerous animals in North America, Europe, and Australia. 1,6,12,15,18 It has also been identified in a central netted dragon (Ctenophorus nuchalis) in northern Australia.6 Partial AgAdV-1 hexon gene sequences in bearded dragons in the United States demonstrated 4 different genotypes; diversity was not seen in the polymerase gene.18 Only 1 other adenovirus has been reported from the genus Pogona, a helodermatid AdV-2-like virus that was identified in a P. minor. 6 Although there have been reports of subclinical adenoviral infection in reptiles, 8,16 many infections result in moderate to severe clinical disease. 576564V DIXXX10.1177/1040638715576564Development of a novel qPCR for AgAdV-1 in bearded dragonsFredholm et al. research-article2015From the Department of Small Animal Clinical Sciences, College of Veterinary Medicine, University of Florida, Gainesville, FL. Abstract. Agamid adenovirus 1 (AgAdv-1) is a significant cause of disease in bearded dragons (Pogona sp.). Clinical manifestations of AgAdv-1 infection are variable and often nonspecific; the manifestations range from lethargy, weigh...

Adenoviruses in free-ranging Australian bearded dragons (Pogona spp.)

“…We decided to screen each dragon using two different PCRs that were capable to detecting adenoviruses in bearded dragons. The PCR described by Fredholm et al (2015) was designed to specifically detect agamid adenovirus-1, which is the most commonly reported adenovirus infection of bearded dragons (Kübber-Heiss et al, 2006;Abbas et al, 2012;Kubiak, 2013;Doneley et al, 2014;Fredholm et al, 2015;Schilliger et al, 2016). In the study by Fredholm et al (2015), this PCR was shown to be able to detect this virus in more samples than the pan-adenovirus PCR described by Wellehan et al (2004).…”

“…The PCR described by Fredholm et al (2015) was designed to specifically detect agamid adenovirus-1, which is the most commonly reported adenovirus infection of bearded dragons (Kübber-Heiss et al, 2006;Abbas et al, 2012;Kubiak, 2013;Doneley et al, 2014;Fredholm et al, 2015;Schilliger et al, 2016). In the study by Fredholm et al (2015), this PCR was shown to be able to detect this virus in more samples than the pan-adenovirus PCR described by Wellehan et al (2004). This result is consistent with our results, where of the nine animals we identified to be infected with agamid adenovirus-1, three were PCR-positive using the PCR by Fredholm et al (2015) but were PCR-negative using the PCR by Wellehan et al (2004).…”

“…Hepatic necrosis, intranuclear inclusion bodies in the liver and gut, and secondary infections or infestations with coccidia and nematodes have been reported (Jacobson, 2007;Hyndman and Shilton, 2011;Doneley et al, 2014;Schilliger et al, 2016). Atadenovirus infections have been described in testudines (Garcia-Morante et al, 2016) and a number of snake and lizard species (Marschang et al, 2003;Farkas et al, 2008;Garner et al, 2008;Abbas et al, 2011;Schmidt et al, 2013), but the most commonly reported host of atadenoviruses are bearded dragons (Moormann et al, 2009;Hyndman and Shilton, 2011;Crossland et al, 2018), with many of the infections being a virus named agamid adenovirus-1 (Kübber-Heiss et al, 2006;Abbas et al, 2012;Kubiak, 2013;Doneley et al, 2014;Fredholm et al, 2015;Schilliger et al, 2016). This higher reporting frequency in bearded dragons is likely contributed to by their popularity as pets.…”

“…Extracted DNA was tested for the presence of adenoviruses using two PCRs: one that was designed to detect all known adenoviruses (Wellehan et al, 2004), and a second one that was designed to specifically detect agamid adenovirus-1 (Fredholm et al, 2015). For the nested PCR described by Wellehan et al (2004), 10 µL of 2x master mix from the Platinum TM Green Hot Start PCR Master Mix (2X) (cat.…”

“…For PCR testing for agamid adenovirus-1, the methods described by Fredholm et al (2015) were followed.…”